The primary goal of this project is to develop a sensor for use in drug development that mimics human brain receptors. This sensor will be capable of evaluating the binding of large numbers of drug candidates in a short period of time;thus greatly enhancing the rate at which potential therapeutic agents can be identified. The primary goal of this project is to develop improved microcantilever-based biosensors using receptor proteins based on a recently discovered acetylcholine binding protein (AChBP). The AChBP is a soluble protein that displays structure and pharmacology that is strikingly similar to the nicotinic acetylcholine receptor (nAChR) present in the central and peripheral nervous systems. Since the nAChR is a member of a large super-family of ligand gated ion channels (LGICs), this project will ultimately develop sensors that will be useful in high throughput drug screening and drug discovery for this class of neuronal receptors. Specifically, we aim to: 1) Develop a microcantilever biosensor utilizing the AChBP and its derivatives as biological sensor proteins. 2) Investigate fundamental surface conjugation chemistries and establish modification protocols for microcantilever sensing. The project draws heavily on our combined expertise in LGIC structure and function (Dr. Schulte) and microcantilever design (Dr. Ji). Aim 1 will evaluate the AChBP itself as a potentially useful biosensor molecule. We will initially demonstrate the use of this sensor in high-throughput drug screening by exploring its interaction with nAChR, GABAR, glycineR and 5-HT3R ligands. Since the AChBP can be mutated to alter its ligand specificity, derivatives obtained via site directed mutagenesis and/or chimeric construction will also be developed. Since the AChBP is commonly used as a template for modeling of ligand gated ion channel receptors, an additional benefit of this work will be an expanded knowledge of the basis of ligand specificity at these important receptors. Aim 2 will utilize the AChBP and other receptor proteins to improve sensor response through improved conjugation chemistries in combination with improved microcantilever array technology developed in Dr. Ji's laboratory